Human cancers are represented by heterogeneous collections of malignant cells that must be eradicated for treatment to be successful. For many solid organ tumors, cancer stem cells (CSCs) are thought to be responsible for treatment failure and poor clinical outcomes. Accordingly, the detection of CSCs represents a significant priority with regard to prognostication, choice of treatment, and for assessing patient responses to inten/ention. Convenfional tools for the detecfion and isolation of CSCs are near exclusively protein limited. Intracellular mRNA targets have been used extensively for cancer cell sub-population phenotyping; however, detecfion of mRNA targets requires destruction of precious candidate cells for performing RT-PCR and sacrifices throughput. Either for basic science research or clinical use, the capability of simultaneous detecfion of both protein and mRNA markers in live cancer cell populafions, and in real fime, would be significantly enabling. This project aims to bring a new and enabling technology, 'NanoFlares', to bear on the detection and isolation of phenotypically distinct cancer stem cells (CSC). Taking advantage of the unique properties of gold nanoparticles surface functionalized with DNA (DNA Au-NPs), the NanoFlare technology provides the unique capability of phenotyping cell sub-populations simultaneously at the protein and mRNA level. Single-cell analysis is possible using confocal light microscopy while thousands of individual cell measurements can take place in high throughput using flow cytometry (FC) and fiuorescence activated cell sorting (FACS). Focusing initially on breast cancer, project success will have significant impact from the standpoint of basic science discovery in the context of CSCs and, ultimately, for pafients with all forms of localized and disseminated cancer where CSC detection and eradication could have a dramatic impact on patient outcomes.